The objective of this research is to develop efficient and rapid procedures for synthsizing polynucleotides possessing any desired sequence of nucleotides. The strategy is to employ three chemical coupling methods that have been explored in this laboratory to link nucleotides and oligonucleotides at different stages in the synthesis. For the stepwise construction of protected oligonucleotides in the six to eight nucleotide range, the phosphite-triester coupling will be used. This procedure is rapid and efficient and especially well suited for syntheses involving addition of one nucleotide unit at a time. For coupling protected oligonucleotides in the six to eight nucleotide range, the phosphotriester synthesis (involving condensation of a phosphodiester with a terminal 5' -OH group) will be employed. Although slower than the phosphite-triester procedure, it affords good yields of coupling products from approximately equimolar amounts of protected oligonucleotides. Finally, for the coupling of long chain oligomers, either in aqueous or nonaqueous media, coupling via internucleotide phosphoramidate bonds will be studied. The basic chemistry for joining small fragments by this method has been worked out and a sound rationale for this approach has been established by the enzymatic synthesis of phosphoramidate DNA. Various types of primer molecules will be made and compared for priming activity in enzymatic extension on a single strand DNA template. A goal of this research is to provide methods for preparing materials that will be useful in biochemical and biological studies aimed at understanding and controlling cancer and genetic diseases.